Glial Cells In Rats Research Articles

The Influence of Aerobic Exercise on Hippocampal Glial Cells and Inflammatory Factor Expression in the Aging Process of Rats

The objective is to analyze the impact of aerobic exercise on the activation of hippocampal glial cells and the expression of inflammatory factors in the aging process of rats. Methods: Thirty male SPF (Specific Pathogen Free) rats at 10 weeks of age were selected and divided into three groups: the sedentary group (A), the aging group (B), and the aging exercise group (C), with 10 rats in each group. The sedentary group (A) was raised for 6 weeks without exercise or aging intervention; the aging group (B) was continuously injected with D-galactose for 6 weeks without exercise intervention; the aging exercise group (C) was continuously injected with D-galactose for 6 weeks while simultaneously implementing exercise intervention, with 60 minutes of unassisted swimming per day, three times a week (swimming pool size 0.5m x 0.8m, water temperature 35±2°C). All groups were harvested at the end of 6 weeks. Compared with the sedentary group, the GFAP immunohistochemical staining in the hippocampal area of the brain was stronger in both the aging group and the aging exercise group, with more GFAP expression, darker positive reaction color, and higher average fluorescence intensity than the sedentary group; compared with the aging group, the aging exercise group had less GFAP expression, lighter positive reaction color, and lower average fluorescence intensity. All differences were significant (P<0.05). In the comparison of GSH levels in the hippocampal tissue of rats, the aging group had the lowest levels, indicating a decrease in the activation level of glial cells in the hippocampus, while the aging exercise group had higher GSH levels than the aging group, proving that exercise intervention in the aging process can effectively improve the activation level of hippocampal glial cells in rats.

Inhibition of NF-κB ameliorates aberrant retinal glia activation and inflammatory responses in streptozotocin-induced diabetic rats.

To determine the anti-inflammatory effects of IMD-0354, a specific NF-κB blocker, on glial cells in rats with streptozotocin (STZ)-induced diabetic retinopathy (DR). The following four groups of rats were used: control, control + IMD-0354, STZ, and STZ + IMD-0354. After six weeks of STZ injection, diabetic rats and nondiabetic control rats received IMD-0354 (30 mg/kg) or an equal volume of 4% dimethyl sulfoxide (DMSO) in phosphate-buffered saline intraperitoneally for six consecutive weeks. The following four groups of primary rat retinal microglia and Müller cells were used: control (5 mM), control + IMD-0354, high glucose (20 mM), and high glucose + IMD-0354. The effects of IMD-0354 on nuclear factor-κB (NF-κB) activation, oxidative stress strength, expression of inflammatory cytokines and VEGF (vascular endothelial growth factor), activation of glial cells, and apoptosis of neuron cells were evaluated by immunohistochemistry, oxidative stress assays, western blot, enzyme linked immunosorbent assay (ELISA) and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining respectively. Nuclear translocation of NF-κB was markedly increased in diabetic rat retina and high glucose treated glial cells. Systemic administration of IMD-0354 significantly inhibited NF-κB activation in both diabetic rat retina and high glucose treated glial cells, ameliorated oxidative injury, inflammatory responses, VEGF production and glial cell activation, and protected neurons from apoptosis. Our findings indicated that NF-κB activation is acritical step in the abnormal reactivity of glial cells in STZ-induced diabetic rats. Inhibition effect of IMD-0354 on NF-κB activation may represent a promising therapeutic strategy for DR via a variety of mechanisms, including inflammation reduction and glial cells regulation.

Tetrahydropalmatine exerts analgesic effects by promoting apoptosis and inhibiting the activation of glial cells in rats with inflammatory pain.

BackgroundPain is an unpleasant sensory experience that usually plays a protective role. Inflammatory pain is often severe and stubborn, which has a great impact on the quality of life of patients. However, there has been no breakthrough in the treatment strategy and mechanism of inflammatory pain.MethodsThis study investigated the analgesic effect of tetrahydropalmatine (THP) in rats injected with complete Freund's adjuvant (CFA)-induced inflammatory pain. Allodynia and gait analysis of rats were used to evaluate the analgesic effect at different time points before and after operation. THP (2.5, 5, and 10 mg/kg) was administered intraperitoneally once daily for 7 days post Day 3. The expression levels of TNF-α and IL-1β in the spinal cord were measured by enzyme-linked immunosorbent assay. The activation of astrocytes and microglial cells in the spinal cord was tested by western blot before and after THP treatment. The apoptosis of glial cells was tested by flow cytometry after treatment with THP in the primary cultured glial cell model.ResultsCFA treatment induced significant allodynia and caused abnormal gait in rats. Administration of THP at 10 mg/kg significantly alleviated CFA-induced inflammatory pain behaviors. Moreover, CFA-induced activation of glial cells and the increased levels of TNF-α and IL-1β were inhibited by THP administration. In addition, THP promotes apoptosis in primary cultured glial cells. This study suggests the possible clinical utility of THP in the treatment of inflammatory pain.ConclusionTHP plays an analgesic role by inhibiting the activation of glial cells and promoting apoptosis.

Transcutaneous Electrical Nerve Stimulation Reduces Knee Osteoarthritic Pain by Inhibiting Spinal Glial Cells in Rats

Transcutaneous electrical nerve stimulation (TENS) is commonly used for pain control. However, the effects of TENS on osteoarthritis (OA) pain and potential underlying mechanisms remain unclear. The objective of this study was to investigate the effect of TENS on OA pain treatment and underlying mechanisms related to glial cell inhibition. This was an experimental study. OA was induced by injection of monosodium iodoacetate into the synovial space of the right knee joint of rats. High-frequency (HF) TENS (100 Hz), low-frequency (LF) TENS (4 Hz), or sham TENS was applied to the ipsilateral knee joint for 20 minutes. Paw withdrawal threshold (PWT), weight bearing, and knee bend score (KBS) were measured. Immunohistochemistry for microglia and astrocytes was performed with L3 to L5 spinal segment samples. To investigate the effects of glial inhibition on OA pain, minocycline, l-α-aminoadipate, or artificial cerebrospinal fluid was injected intrathecally, and PWT and KBS were measured. Compared with sham TENS, both HF TENS and LF TENS significantly increased PWT, decreased KBS, and inhibited activated microglia in the L3 to L5 segments but did not decrease the total number of microglia, except in the L4 segment (HF TENS). Astrocyte expression was significantly decreased in the L3 to L5 segments following LF TENS and in the L3 segment following HF TENS. Compared with artificial cerebrospinal fluid, both minocycline and l-α-aminoadipate increased PWT and decreased KBS. These results cannot be generalized to humans. TENS alleviates OA pain in rats by inhibiting activated microglia and reducing astrocyte expression in the spinal cord. Although these results may not be generalizable to chronic pain in patients with OA, within the limitation of the experimental animal model used in the present study, they suggest a possible mechanism and preclinical evidence supporting further experimentation or clinical use of TENS in humans.

Oxycodone ameliorates the inflammatory response induced by lipopolysaccharide in primary microglia.

BackgroundActivation of microglia participates in a wide range of pathophysiological processes in the central nervous system. Some studies reported that oxycodone (6-deoxy-7,8-dehydro-14-hydroxy-3-O-methyl-6oxomorphine) could inhibit the overactivation of glial cells in rats’ spinal cords. In the present study, we observed the effect of oxycodone on inflammatory molecules and pathway in lipopolysaccharide (LPS)-stimulated primary microglia in rats.Materials and methodsNeonatal rats’ primary microglia were exposed to various concentrations (25, 50, 100 ng/mL) of oxycodone for 1 h after LPS stimulation for 24 h. The levels of pro-inflammatory mediators, IL-1β, TNF-α, and TGF-β1/smad2/3 signaling pathway were measured. The activation situation of microglia and the expression of TβR1 were observed by immunofluorescence.ResultsOxycodone at 25 ng/mL did not change the levels of proinflammatory molecules and TGF-β1/smad2/3 signaling pathway in primary microglia, which was increased by LPS. Oxycodone at 50 and 100 ng/mL could significantly suppress LPS-induced production of TNF-α and IL-1β and the expression of TNF-αmRNA, IL-1βmRNA, and TGF-β1/smad2/3 signaling pathway.ConclusionThese findings indicate that oxycodone, at relatively high clinically relevant concentration, can inhibit inflammatory response in LPS-induced primary microglia. The detailed mechanism needs to be investigated in future.

A Single Intraperitoneal Injection of Endotoxin Changes Glial Cells in Rats as Revealed by Positron Emission Tomography Using [11C]PK11195.

Intracranial administration of lipopolysaccharide (LPS) is known to elicit a rapid innate immune response, activate glial cells in the brain, and induce depression-like behavior. However, no study has focused on the changes in glial cells induced by intraperitoneal injection of LPS in vivo. Ten adult male Fischer F344 rats underwent [11C]PK11195 PET before and 2days after intraperitoneal injection of LPS to evaluate the changes in glial cells. The difference in standardized uptake values (SUV) of [11C]PK11195 between before and after injection was determined. There was a cluster of brain regions that showed significant reductions in SUV. This cluster included the bilateral striata and bilateral frontal regions, especially the somatosensory areas. Changes in activity of glial cells induced by the intraperitoneal injection of LPS were detected in vivo by [11C]PK11195 PET. Intraperitoneal injection of LPS is known to induce depression, and further studies with [11C]PK11195 PET would clarify the relationships between neuroinflammation and depression.

The effects of lower thoracic epidural blockade on intestinal epithelial barrier and activation of enteric glial cells in rats with hemorrhagic shock and resuscitation

Objective To investigate the effects of lower thoracic epidural blockade on integrity of intestinal epithelial barrier(IEB) and activation of enteric glial cell(EGC) in rats with hemorrhagic shock and resuscitation. Methods Seventy-two SD rats with epidural catheterization according to a modified method described by Bahar et al. were randomly divided into four groups with 18 in each group: sham group(group S), control group(group C), normal saline group(group N) and thoracic epidural blockade group(group T). In group S, rats were just received vascular cannulation, whereas the rats in other groups were subjected to hemorrhagic shock and resuscitation with normal saline according to the modified method described by Wiggers. Thirty minutes before bleeding, either 0.075% ropivacaine(group T) or normal saline(group N) 100 μl/kg was infused through thoracic epidural catheters. The survival rates of rats in 72 h after resuscitation were evaluated. The MAP and HR were monitored after vascular cannulation(T0), 30 min after epidural infusion(T1), 30 min of shock(T2), 60 min of shock(T3), 30 min and 60 min after resuscitation(T4 and T5 respectively). Arterial blood samples were collected for blood gas analysis at T0, T3 and T5. At 3 h following resuscitation, the severity of intestinal edema, the pathological changes of intestinal mucosa, and the function of IEB were evaluated by wet/dry ratio(W/D), Chiu's score, and lipopolysaccharide(LPS) concentrations in the portal vein, respectively. The expression of Occludin and glial fibrillary acidic protein(GFAP), a marker of EGCs, were determined by western blot. Results Compared with group C and group N, the survival rates and median survival time of group T were significantly promoted (P<0.05), While MAP and HR decreased at T4 and T5, the levels of pH and lactic acid at T3 and T5 were alleviated (P<0.05). Besides these, in group T, the levels of W/D, Chiu's score and LPS were decreased while the expression of Occludin and GFAP were increased significantly in comparison with group C and group N(P<0.05). Conclusions Lower thoracic epidural blockade attenuates IEB impairment induced by hemorrhagic shock and resuscitation in rats, alleviating internal environment disturbance and thus promoting survival rates after resuscitation, whose mechanism is related to activation of EGC. Key words: Thoracic epidural blockade; Hemorrhagic shock; Resuscitation; Intestinal epithelial barrier; Enteric glial cell

Maternal High Fat Diet Induces Myenteric Neuronal Loss and Upregulates Myenteric Glia Prior the Development of Obesity or Inflammation

Diet‐induced obesity disrupts gastrointestinal (GI) functions possibly due to a loss of myenteric neurons, although few studies have assessed the effects of diet on the development of these neurons. The aim of the present study was to assess the effects of exposure to a high fat diet (HFD) during the development period on myenteric neurons and glial cells in rats and the possible role of systemic and local inflammation.Sprague‐Dawley rats were fed a control or HFD (14% or 60% kcal from fat, respectively) from embryonic day 13; the fundus, corpus and duodenum were removed and fixed at postnatal ages 2, 4, 6 and 12 weeks (n=4 for each timepoint). Tissues were processed for immunohistochemical detection of total myenteric neurons (protein gene product 9.5; PGP9.5) as well as cholinergic (choline acetyltransferase; ChAT) and nitrergic (nitric oxide synthase; NOS) neurons, RNA‐binding protein (HuC/D) and glial cells (glial fibrillary acid protein; GFAP). Confocal microscope images were analyzed using Image J software. Serum and gastrointestinal levels of the cytokines IL‐1β and IL‐6 were quantified via multiplex and qRT‐PCR, respectively.HFD rats were overweight but not regarded as obese until 12 weeks of age (292±12g vs 380±12g). By 4 weeks of age, however, a decrease in number of myenteric neurons per ganglia was observed in the fundus (29 ± 3.6 vs. 38 ± 2.6 neurons/ganglia; P&lt;0.05) and corpus (34.6 ± 0.8 vs. 44.3 ± 1.2 neurons/ganglia; P&lt;0.05), and by 6 weeks of age in the duodenum (26 ± 1.5 vs. 34.6 ± 2.9 neurons/ganglia; P&lt;0.05). A selective loss of nitrergic neurons was observed by 12 weeks of age in the fundus (13.5 ± 0.9% vs 20 ± 0.9% of total neurons; P&lt;0.05), corpus (10.7 ± 0.7% vs 19 ± 3.1%; P&lt;0.05), and duodenum (8.5 ± 2.5% vs 16.5 ± 2.0%; P&lt;0.05). HFD exposure increased myenteric glial cell density by at 4 weeks of age in the all regions studied (fundus 69.9±2.6 vs 62.32±3.1 pixels/cm2; corpus 87.0±7.5 vs 62.9±4.9 pixels/cm2; and duodenum (96.1±5.9 vs 56.9±4.0 pixels/cm2; P&lt;0.05 for each), suggesting a possible diet‐induced inflammation of the gastrointestinal tract. While nuclear aggregation of HuC/D, suggesting compromised neuronal health, was observed in myenteric neurons at 12 weeks of age in all regions studied (fundus 68.6 ± 8.5% vs 41.6 ± 6.6% nuclear localization; corpus 55.0 ± 5.4% vs 23.5 ± 3.4%; and duodenum 52.4 ± 5.5% vs 34.7 ± 1.0%; P&lt;0.05 for each), there were no consistent alterations in serum or gastrointestinal expression of IL‐1β (22.98±2.73 vs 28.57±2.03 pg/mL and 0.67±0.19 vs 0.32±0.09 2DDCT, respectively; P&gt;0.05) and IL‐6 (88±10.17 vs 115±7.34 pg/mL; 0.94±0.39 vs 1.00±0.34 vs ± 2DDCT;P&gt;0.05).The present study suggests that exposure of rats to a HFD from the developmental period induces loss of inhibitory nitrergic neurons accompanied by glial proliferation even prior to the onset of obesity and in the absence of elevated cytokine levels, suggesting that diet may alter gastrointestinal functions independently of increased adiposity or adipose‐related inflammation.Support or Funding InformationFunded by NIH 078364 and NSF IOS1148978

Effects of glial cell derived neurotrophic factor on colon glial cells in rats with slow transit constipation

Objective To investigate the effects of exogenous glial cell derived neurotrophic factor (GDNF) on colon glial cells in slow transit constipation (STC) rats, and to explore the optimal concentration of GDNF in order to provide evidence for intestinal neurotrophic therapy in the treatment of STC. Methods A total of 132 SD rats were divided into STC group and control group, 66 rats in each group. STC rats were established by feeding with rhubarb. Six rats were randomly selected from either groups to verify whether STC model was successfully established.And the left 120 rats of two groups were randomly divided into six subgroups: STC group one to group six and control group one to group six, ten rats in each group, which were untreated, injected through tail vein with saline, and 0.001, 0.010, 0.050, 0.100 μg/L GDNF 2 mL respectively for one week. The expression of Sox-8 at protein level of either group were detected by Western blotting. Independent sample t test was performed for statistical analysis. Results After treated with 0.001 μg/L GDNF (STC group three), there was no significant difference in expression level of Sox-8 between STC group three and STC group one (13.38±0.70 vs 13.39±0.45, t=0.042, P=0.969). After treated with 0.010 μg/L GDNF (STC group four), the difference in expression level of Sox-8 between STC group four and STC group three was significant (21.11±2.56 vs 13.38±0.70, t=5.040, P<0.01). After treated with 0.050 μg/mL GDNF (STC group five), the expression level of Sox-8 was higher than that in STC group four (31.86±1.57 vs 21.11±2.56, t=-6.198, P<0.01). The Sox-8 expression of untreated, saline treated, 0.001 and 0.050 μg/L GDNF treated STC rats (STC group one, two, three and five) were lower than those of the corresponding control groups (t=3.394, 12.103, 10.302, -6.120, all P<0.05). Conclusion Exogenous GDNF could increase Sox-8 expression in colon tissue of STC rats, an increase in the number of colon glial cells could repair enteric nervous system, and 0.050 μg/L was the optimal concentration. Key words: Slow transit constipation; Glial cell line-derived neurotrophic factor; Colon glial cell; Sox-8; Rats

Synchronized dual pulse gastric electrical stimulation induces activation of enteric glial cells in rats with diabetic gastroparesis.

Objective. The aims of this study were to investigate the effects of synchronized dual pulse gastric electrical stimulation (SGES) on gastric motility in different periods for diabetic rats and try to explore the possible mechanisms of the effects. Methods. Forty-six rats were used in the study. Gastric slow waves were recorded at baseline, 7–14-day diabetes and 56–63-day diabetes before and after stimulation and the age-matched control groups. SGES-60 mins and SGES-7 days (60 mins/day) were performed to test the effects on gastric motility and to evaluate glial marker S100B expression in stomach. Results. (1) Gastric emptying was accelerated in 7–14-day diabetes and delayed in 56–63-day diabetes. (2) The S100B expression in 56–63-day diabetes decreased and the ultrastructure changed. (3) The age-associated loss of EGC was observed in 56–63-day control group. (4) SGES was able to not only accelerate gastric emptying but also normalize gastric slow waves. (5) The S100B expression increased after SGES and the ultrastructure of EGC was partially restored. The effect of SGES-7 days was superior to SGES-60 mins. Conclusions. Delayed gastric emptying due to the growth of age may be related to the EGC inactivation. The effects of the SGES on gastric motility may be associated with EGC activation.

Involvement of trigeminal ganglionic Nav1.7 in hyperalgesia of inflamed temporomandibular joint is dependent on ERK1/2 phosphorylation of glial cells in rats

Inflammation is a major cause of temporomandibular disorder-related pain. The Nav 1.7 sodium channel has a critical function in pain perceptions. However, whether and how Nav 1.7 in the trigeminal ganglion is involved in temporomandibular joint (TMJ) inflammatory pain remains to be examined. TMJ inflammation was induced by complete Freund's adjuvant in female rats. The expression of trigeminal ganglionic Nav 1.7 and other sodium channels was examined using real-time polymerase chain reaction or Western blotting. Immunohistofluorescence with fluorescent retrograde neuronal tracer DiI was used to confirm Nav 1.7 in the trigeminal neurons innervating TMJ. The functions of trigeminal ganglionic Nav 1.7 and extracellular signal-regulated kinase 1/2 (ERK1/2) phosphorylation were blocked with the microinjection of the Nav 1.7 antibody or U0126 into the trigeminal ganglion. Head withdrawal threshold and food intake was measured to evaluate TMJ nociceptive responses. TMJ inflammation significantly up-regulated Nav 1.7 mRNA and protein; however, the mRNA of Nav 1.3 was not affected and those of Nav 1.8 and Nav 1.9 were only slightly up-regulated. TMJ inflammation specifically induced Nav 1.7 in the neurons innervating TMJ. In addition, blocking the Nav 1.7 function significantly attenuated the hyperalgesia of the inflamed TMJ. Moreover, TMJ inflammation up-regulated ERK1/2 phosphorylation only in the glials; blocking ERK1/2 phosphorylation in the glials blocked Nav 1.7 up-regulation in the neurons and correspondingly attenuated the hyperalgesia of the inflamed TMJ. Trigeminal ganglionic Nav 1.7 has an important function in the hyperalgesia of the inflamed TMJ, which is dependent on the communication with the satellite glials.

The effects of centrally administered fluorocitrate via inhibiting glial cells on working memory in rats

Although prefrontal and hippocampal neurons are critical for spatial working memory, the function of glial cells in spatial working memory remains uncertain. In this study we investigated the function of glial cells in rats' working memory. The glial cells of rat brain were inhibited by intracerebroventricular (icv) injection of fluorocitrate (FC). The effects of FC on the glial cells were examined by using electroencephalogram (EEG) recordings and delayed spatial alternation tasks. After icv injection of 10 microL of 0.5 nmol/L or 5 nmol/L FC, the EEG power spectrum recorded from the hippocampus increased, but the power spectrum for the prefrontal cortex did not change, and working memory was unaffected. Following an icv injection of 10 microL of 20 nmol/L FC, the EEG power spectra in both the prefrontal cortex and the hippocampus increased, and working memory improved. The icv injection of 10 microL of 50 nmol/L FC, the EEG power spectra in both the prefrontal cortex and in the hippocampus decreased, and working memory was impaired. These results suggest that spatial working memory is affected by centrally administered FC, but only if there are changes in the EEG power spectrum in the prefrontal cortex. Presumably, the prefrontal glial cells relate to the working memory.

Reactive changes of retinal astrocytes and Müller glial cells in kainate‐induced neuroexcitotoxicity

The aim of this study was to investigate reactive changes of astrocytes and Müller glial cells in rats subjected to kainate treatment, which leads to neuronal degeneration in the ganglion cell layer and the inner border of the inner nuclear layer as confirmed by labelling with Fluoro-Jade B, a marker for degenerating neurons and fibres. Both the astrocytes and the Müller glial cells reacted vigorously to kainate injection as shown by their up-regulated expression of nestin, glial fibrillary acidic protein and glutamine synthetase. A major finding was the induced expression of nestin together with glial fibrillary acidic protein beginning at 1 day post-injection of kainate. The marked nestin expression appeared to be most intense at 1 day and was sustained till 2 weeks as compared with the untreated/normal retina. Western blotting analysis confirmed a marked increase in expression of nestin, glial fibrillary acidic protein and glutamine synthetase as compared with untreated/normal retina. Double labelling study revealed that astrocytes and Müller glial cells expressed the radial glia marker nestin, and incorporated bromodeoxyuridine to re-enter into their cell cycle. The induced expression of these proteins in astrocytes and Müller glial cells indicated an induction of gliotic responses and de-differentiation that may be associated with regenerative efforts after kainate-induced injury. Indeed, with the acquisition of an immature molecular profile as manifested by the induced expression of brain lipid-binding protein and doublecortin in astrocytes and Müller glial cells, the potential of these cells to de-differentiate in retinal neurodegeneration is greatly amplified.

Diabetes Alters Osmotic Swelling Characteristics and Membrane Conductance of Glial Cells in Rat Retina

The development of edema in the diabetic retina may be caused by vascular leakage and glial cell swelling. To determine whether diabetic retinopathy alters the swelling characteristics of retinal glial cells and changes the properties of the glial membrane K+ conductance, isolated retinas and glial cells of rats were investigated at 4 and 6 months of chemical diabetes. After 6 months of hyperglycemia, application of a hypotonic solution to retinal slices induced swelling of glial cell bodies, a response not observed in control retinas. The osmotic glial cell swelling was blocked by inhibitors of phospholipase A2 or cyclooxygenase and by a thiol-reducing agent. Glial cells from diabetic retinas displayed a decrease of K+ currents that was associated with an altered subcellular distribution of the K+ conductance and a loss of perivascular Kir4.1 protein. The observation that swelling of cells in control retinas was inducible with K+ channel-blocking Ba2+ ions suggests a relationship between decreased K+ inward currents and osmotic cell swelling in diabetic retinas. The data show that glial cells in diabetic retinas are more sensitive to osmotic stress, which is associated with a decrease of K+ currents, than cells in control retinas. It is suggested that these alterations may be implicated in the development of diabetic retinal edema.

Müller glial cells express nestin coupled with glial fibrillary acidic protein in experimentally induced glaucoma in the rat retina

This study was aimed to investigate reactive changes of Müller glial cells in rats subjected to experimentally induced glaucoma. In the latter, it is well documented that elevated intraocular pressure leads to the loss of ganglion cells as confirmed in this study. The present results have shown that Müller glial cells as well as astrocytes closely associated with the ganglion cells reacted vigorously to increased intraocular pressure as manifested by the induced and upregulated expression of nestin and glial fibrillar acidic protein. A major finding in glaucomatous rats was the induced expression of nestin together with glial fibrillar acidic protein with the rise of the intraocular pressure beginning at 2 h. The marked nestin expression appeared to be most intense at 1 week after operation and was sustained at 3 weeks. Induced nestin expression in Müller glial cells was demonstrated unequivocally in whole-mount preparation of the retina. In the same tissue preparation, nestin expression was also detected in some astrocytes. Western blotting analysis confirmed a marked increase in expression of nestin and glial fibrillar acidic protein. Present results suggest that nestin as well as glial fibrillar acidic protein is a useful biomarker for retina injury. The induced expression of these intermediate filament proteins in Müller glial cells especially at their end-feet and also in some astrocytes adjoining the neuronal injury suggests a potential neuroprotective mechanism in response to acute rise in intraocular pressure resulting in neuronal degeneration.

Small Changes of Dietary (n-6) and (n-3)/Fatty Acid Content Ratio Alter Phosphatidylethanolamine and Phosphatidylcholine Fatty Acid Composition During Development of Neuronal and Glial Cells in Rats

It has been suggested that the fat composition of infant formula should provide arachidonic acid [20:4(n-6)] and docosahexaenoic acid [22:6(n-3)] or increased α-linolenic acid [18:3(n-3)] to optimize the (n-3) and (n-6) fatty acid content of brain during infant development. This experiment examined the effects of feeding increased levels of 18:3(n-3), 20:4(n-6) and 22:6(n-3) on brain development in neonatal rats. Diets varying in (n-6) and (n-3) fatty acid content with or without 20:4(n-6) or 22:6(n-3), at levels proposed for infant formula, were fed to nursing dams from parturition and subsequently to weaned pups until 6 wk of age. Neuronal and glial cells were isolated from the frontal region, cerebellum and hippocampus of the brain. Fatty acid analyses of ethanolamine- and choline-phosphoglycerides indicated that small changes in the dietary (n-6)/(n-3) ratio significantly altered neuronal and glial membrane fatty acid composition. Brain regions and cell types varied in amount and rate of 20:4(n-6) and 22:6(n-3) accretion. Fatty acid composition of individual phosphoglycerides was distinct and exhibited changes with age. Inclusion of both 20:4(n-6) and 22:6(n-3) in the diet resulted in alteration of brain fatty acid composition reflecting the fatty acid composition of the diet. If analogous developmental changes occur in human brain, then these results imply that addition of 20:4(n-6) and 22:6(n-3) or a reduced 18:2(n-6):18:3(n-3) ratio in infant formula may result in fatty acid profiles of neuronal and glial cells in formula-fed infants similar to those observed in breast-fed infants.

High resolution AMS imaging of radiocarbon in biomedical applications

Radiocarbon has been an important labelling element in biological metabolism studies. By interfacing an accelerator mass spectrometer (AMS) with a scanning microprobe secondary ion source, we have imaged the uptake of radiocarbon labelled metabolic or neurotransmitter amino acids by neurons and glial cells of rats and gerbils at high resolution (1 micron), high sensitivity and in a short time. The biological samples are prepared and sectioned serially at 0.5 μm thickness using standard histological procedures. The adjacent sections to those used for AMS imaging were either immunolabelled with antibodies to GABA to reveal GABA-containing cells, or stained with toluidine blue to visualise every cell. Therefore, the distribution of radiocarbon revealed by AMS could be matched to that of the cells. By simultaneously measuring the 14C, 13C and 12C signals, we can demonstrate that the localised peaks of radiocarbon could be readily identified and matched to GABA-immunopositive neurons and glial cells by aligning the radiocarbon deficient blood vessels with the vessels in the adjacent histologically stained section. The results revealed the selective uptake of the neurotransmitter, GABA and that of metabolic amino acid, leucine. The technique compares favourably with high resolution autoradiography and provides great potential for improving the analysis of molecular interactions in and between cells.

Protein synthesis in neurons and satellite glial cells of rats after global cerebral ischemia caused by cardiac arrest

Protein synthesis rates are increased in light and dark neurons and satellite glia cells in various brain areas in rats with completely restituted neurological status after cardiac arrest. However, the increase is provided by different cell types. Only free light neurons are reactive in the hippocampal sector CA1, while both free light neurons as light neurons with glia are reactive in sector CA4; intense protein synthesis is observed in all types of the cerebral cortex neurons and in cortical neurons and in the satellite glial cells of the lateral cerebellar hemispheres.